AUTHOR=Wang Yi , Yan Weiqiang , Fu Shanshan , Hu Shoukui , Wang Yan , Xu Jianguo , Ye Changyun 

TITLE=Multiple Cross Displacement Amplification Coupled With Nanoparticles-Based Lateral Flow Biosensor for Detection of Staphylococcus aureus and Identification of Methicillin-Resistant S. aureus

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.00907

DOI=10.3389/fmicb.2018.00907

ISSN=1664-302X

ABSTRACT=<p><italic>Staphylococcus aureus</italic> (<italic>S. aureus</italic>), including methicillin-resistant <italic>S. aureus</italic> (MRSA), is one of the most important human pathogens, which is responsible for bacteremia, soft-tissue infections, and food poisoning. Hence, multiple cross displacement amplification (MCDA) is employed to detect all <italic>S. aureus</italic> strains, and differentiates MRSA from methicillin-sensitive <italic>S. aureus</italic>. Multiplex MCDA (m-MCDA), which targets the <italic>nuc</italic> gene (<italic>S. aureus</italic>-specific gene) and <italic>mecA</italic> gene (encoding penicillin-binding protein-2′), could detect <italic>S. aureus</italic> strains and identify MRSA within 85 min. Detection of the m-MCDA products is achieved using disposable lateral flow biosensors. A total of 58 strains, including various species of Gram-positive and Gram-negative strains, are used for evaluating and optimizing m-MCDA assays. The optimal amplification condition is found to be 63°C for 40 min, with detection limits at 100 fg DNA/reaction for <italic>nuc</italic> and <italic>mecA</italic> genes in the pure cultures, and 10 CFU/tube for <italic>nuc</italic> and <italic>mecA</italic> genes in the blood samples. The analytical specificity of m-MCDA assay is of 100%, and no cross-reactions to non-<italic>S. aureus</italic> strains are produced according to the specificity testing. Particularly, two additional components, including AUDG enzyme and dUTP, are added into the m-MCDA amplification mixtures, which are used for eliminating the unwanted results arising from carryover contamination. Thus, the m-MCDA technique appears to be a simple, rapid, sensitive, and reliable assay to detect all <italic>S. aureus</italic> strains, and identify MRSA infection for appropriate antibiotic therapy.</p>